Treatment for diseases involving inflammation

ABSTRACT

The present invention relates to a method to protect an animal from a disease involving inflammation by treating that animal with an effective amount of IL-12. The present invention also relates to a method for prescribing treatment for a respiratory disease involving an inflammatory response and a method for monitoring the success of a treatment for a respiratory disease involving an inflammatory response in an animal. Also included in the present invention is a formulation comprising IL-12 and a compound capable of enhancing the effectiveness of the IL-12 at protecting an animal from a disease involving inflammation.

This invention was made in part with government support under POI 36577awarded by the National Institutes of Health. The government has certainrights to this invention.

FIELD OF THE INVENTION

The present invention is related to a method to protect an animal from adisease involving inflammation, in particular, a respiratory diseaseinvolving inflammation.

BACKGROUND OF THE INVENTION

Diseases involving inflammation are characterized by the influx ofcertain cell types and mediators, the presence of which can lead totissue damage and sometimes death. Diseases involving inflammation areparticularly harmful when they afflict the respiratory system, resultingin obstructed breathing, hypoxemia, hypercapnia and lung tissue damage.Obstructive diseases of the airways are characterized by airflowlimitation (i.e., airflow obstruction or narrowing) due to constrictionof airway smooth muscle, edema and hypersecretion of mucous leading toincreased work in breathing, dyspnea, hypoxemia and hypercapnia. Whilethe mechanical properties of the lungs during obstructed breathing areshared between different types of obstructive airway disease, thepathophysiology can differ.

A variety of inflammatory agents can provoke airflow limitationincluding allergens, cold air, exercise, infections and air pollution.In particular, allergens and other agents in allergic or sensitizedanimals (i.e., antigens and haptens) cause the release of inflammatorymediators that recruit cells involved in inflammation. Such cellsinclude lymphocytes, eosinophils, mast cells, basophils, neutrophils,macrophages, monocytes, fibroblasts and platelets. Inflammation resultsin airway hyperresponsiveness. A variety of studies have linked thedegree, severity and timing of the inflammatory process with the degreeof airway hyperresponsiveness. Thus, a common consequence ofinflammation is airflow limitation and/or airway hyperresponsiveness.

Currently, therapy for treatment of inflammation predominantly involvesthe use of glucocorticosteroids. Other anti-inflammatory agents are usedincluding cromolyn and nedocromil. Symptomatic treatment withbeta-agonists, anticholinergic agents and methyl xanthines areclinically beneficial for the relief of discomfort but fail to stop theunderlying inflammatory processes that cause the disease. The frequentlyused systemic glucocorticosteroids have numerous side effects,including, but not limited to, weight gain, diabetes, hypertension,osteoporosis, cataracts, atherosclerosis, increased susceptibility toinfection, increased lipids and cholesterol, and easy bruising.Aerosolized glucocorticosteroids have fewer side effects but can be lesspotent and have side effects, such as thrush.

Other anti-inflammatory agents, such as cromolyn and nedocromil are muchless potent and have fewer side effects. Anti-inflammatory agents thatare primarily used as immunosuppressive agents and anti-cancer agents(i.e., cytoxan, methotrexate and Immuran) have also been used to treatinflammation. These agents, however, have serious side effect potential,including, but not limited to, increased susceptibility to infection,liver toxicity, drug-induced lung disease, and bone marrow suppression.Thus, such drugs have found limited clinical use for the treatment ofmost airway hyperresponsiveness lung diseases.

The use of anti-inflammatory and symptomatic relief reagents is aserious problem because of their side effects or their failure to attackthe underlying cause of an inflammatory response. There is a continuingrequirement for less harmful and more effective reagents for treatinginflammation. Thus, there remains a need for processes using natural(i.e., non-synthetic) reagents with lower side effect profiles and lesstoxicity than current anti-inflammatory therapies.

SUMMARY OF THE INVENTION

The present invention provides for a method and a formulation forprotecting an animal from diseases involving inflammation. The presentinvention is particularly advantageous in that it provides for a the useof a natural substance that is not a synthetically-derived drug, therebyreducing the side effects and toxicity profiles frequently associatedwith anti-inflammatory therapies.

One embodiment of the present invention includes a method to protect ananimal from a disease involving an inflammatory response, the methodcomprising administering to the animal an effective amount ofinterleukin-12. The method of the present invention is particularlyeffective in protecting animals from lung diseases caused byinflammation and skin diseases caused by inflammation. Preferably, theanimal is desensitized to a disease by administering the interleukin-12.In particular, an animal is desensitized against an antigen, anallergen, a hapten, a drug and/or an occupational asthma causing agent.

Another embodiment of the present invention is a method for prescribingtreatment for a respiratory disease involving an inflammatory response,comprising: (1) administering to an animal an effective amount ofinterleukin-12; (2) measuring a change in lung function in response to aprovoking agent in the animal to determine if the interleukin-12 iscapable of modulating airway hyperresponsiveness; and (3) prescribing apharmacological therapy effective to reduce inflammation based upon thechanges in lung function. Preferred provoking agents include direct andindirect stimuli.

Yet another embodiment of the present invention includes a method formonitoring the success of a treatment for a respiratory diseaseinvolving an inflammatory response in an animal, the method comprising:(1) administering an effective amount of interleukin-12 to an animalthat has been treated for a respiratory disease involving aninflammatory response; (2) measuring a change in lung function in theanimal in response to a provoking agent; and (3) monitoring the successof the treatment by comparing the change in lung function with previousmeasurements of lung function in the animal.

The present invention also includes a formulation for protecting ananimal from a disease involving inflammation, comprising interleukin-12and an allergen, an antigen, a hapten, IL-2, IL-4, glucocorticosteroids,anti-cyclooxygenase agents, anti-cholinergic agents, beta-adrenergicagonists, methyl xanthines, anti-histamines, cromones, zyleuton,anti-CD4 reagents, anti-IL-5 reagents, surfactants, anti-thromboxanereagents, anti-serotonin reagents, ketotiphen, cytoxin, cylosporin,methotrexate, macrolide antibiotics, troleadomycin, heparin, and/or lowmolecular weight heparins.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1 illustrates the effect of IL-12 on the respiratory systemfunction of a first group of BALB/c mice challenged with methacholine.

FIG. 2 illustrates the effect of IL-12 on the respiratory systemfunction of a second group of BALB/c mice challenged with methacholine.

FIG. 3 illustrates the combined standard mean effect of IL-12 on therespiratory system function of a first and a second group of BALB/c micechallenged with methacholine.

DETAILED DESCRIPTION OF THE INVENTION

One embodiment of the present invention is a method to protect an animalfrom a disease involving inflammation, comprising administering to theanimal an effective amount of interleukin-12 (IL-12). The inventors havediscovered that IL-12 is a significant inhibitor of inflammation.Previous investigators have reported the use of IL-12 in diseases 5including bacterial, viral and parasitic infections, anemia, T and Bcell deficiencies and tumorigenesis (Trinchieri et al., PCT PublicationNo. WO 92/05256, published Apr. 2, 1992; Trinchieri et al., PCTPublication No. W0 92/05147, published May 17, 1990; Afonso et al., pp.235-236, 1994, Science, Vol. 263, January 14; Sypek et al., pp.1797-1802, 1993, J. Exp. Med., Vol. 177, June; Heinzel et al., pp.1505-1509, 1993, J. Expt. Med., Vol. 177, May; and Hsieh et al., pp.547-549, 1993, Science, Vol. 260, April 23). Such reports, however, donot disclose the use of IL-12 to treat diseases involving inflammationwhich can involve different cellular and humoral responses than thediseases in the reports.

According to the present invention, IL-12 can be administered to anymember of the kingdom Animalia including, without limitation, primates,rodents, livestock and domestic pets. A preferred animal to protectusing IL-12 includes a human, a cat and a horse. A preferred horse toprotect with IL-12 includes a racehorse or a showhorse.

As used herein, the phrase "to protect an animal from a diseaseinvolving inflammation" refers to reducing the potential for aninflammatory response (i.e., a response involving inflammation) to aninflammatory agent (i.e., an agent capable of causing an inflammatoryresponse, e.g., methacholine, histamine, an allergen, a leukotriene,saline, hyperventilation, exercise, sulfur dioxide, adenosine,propranolol, cold air, antigen and bradykinin). Preferably, thepotential for an inflammatory response is reduced, optimally, to anextent that the animal no longer suffers discomfort and/or alteredfunction from exposure to the inflammatory agent. For example,protecting an animal can refer to the ability of a compound, whenadministered to the animal, to prevent a disease from occurring and/orcure or alleviate disease symptoms, signs or causes. In particular,protecting an animal refers to modulating an inflammatory response tosuppress (e.g., reduce, inhibit or block) an overactive or harmfulinflammatory response. Also in particular, protecting an animal refersto regulating cell-mediated immunity and/or humoral immunity (i.e., Tcell activity and/or IgE activity). Disease refers to any deviation fromnormal health of an animal and include disease symptoms as well asconditions in which a deviation (e.g., infection, gene mutation, geneticdefect, etc.) has occurred but symptoms are not yet manifested.

In a preferred embodiment, the present invention protects an animal fromincludes a lung disease caused by inflammation or a skin disease causedby inflammation (e.g., atopic dermatitis). In a more preferredembodiment, the present invention protects an animal from includes achronic obstructive pulmonary disease (COPD) of the airways (i.e.,airway obstruction caused by infiltration of inflammatory cells,scarring, edema, smooth muscle hypertrophy/hyperplasia, smooth musclecontraction and narrowing due to secretions, e.g., mucous, by cells). Inan even more preferred embodiment, the present invention protects ananimal from a physiological impairment, including asthma, allergicbronchopulmonary aspergillosis, hypersensitivity pneumonia, eosinophilicpneumonia, emphysema, bronchitis, allergic bronchitis bronchiectasis,cystic fibrosis, hypersensitivity pneumotitis, occupational asthma(i.e., asthma, wheezing, chest tightness and cough caused by asensitizing agent, such as an allergen, irritant or hapten, in the workplace), sarcoid, reactive airway disease syndrome (i.e., a singleexposure to an agent that leads to asthma), interstitial lung disease,hyper-eosinophilic syndrome or parasitic lung disease. In a preferredembodiment, the present invention protects an animal from asthma,occupational asthma and reactive airway disease syndrome.

Preferably, protecting an animal from a disease involving inflammationincludes desensitizing an animal by administering an effective amount ofIL-12 to the animal. As used herein, desensitizing an animal refersreducing the animal's immune response to a particular compound capableof causing an inflammatory response. Desensitizing an animal against aparticular compound can include immunizing or tolerizing to a particularcompound. Immunization refers to stimulating an immune response by, forexample, activating T helper cells to secrete cytokines which stimulateimmunoglobulin production by B cells. Tolerizing refers to inhibiting animmune response by, for example, killing or anergizing (i.e.,diminishing reactivity by a T cell to an antigenic peptide) particularcells involved in the immune response.

Suitable agents against which to desensitize an animal against includecompounds capable of causing inflammation. Preferred compounds againstwhich to desensitize an animal against include, but are not limited to,an allergen, an antigen, a hapten, a drug and/or an occupational asthmacausing agent. An allergen refers a compound capable of inducing anallergic or hypersensitive response. An antigen refers to a compound(i.e., a foreign substance) that binds specifically to an antibody or aT cell receptor and elicits an immune response. A hapten refers to a lowmolecular weight compound that is not immunogenic by itself, but, whencomplexed or coupled to a larger carrier molecule (i.e., a highmolecular weight protein), can elicit antibodies directed against thehapten. A drug refers to any chemical compound that can be administeredto an animal as an aid in the diagnosis, treatment or prevention ofdisease or an abnormal condition. An occupational asthma causing agentrefers to an agent encountered in a workplace environment that iscapable of causing airway obstruction. Such agents can includeallergens, antigens and haptens. Occupational asthma causing agents thatare haptens include, but are not limited to, benzene derivatives (e.g.,aminobenzenes, aminobenzoic acid), toluidine, toluidine derivatives(e.g., toluidine diisocyanate) metals, amines, acid anhydrides andplicatic acid. Other occupational asthma causing agents include, forexample, cigarette smoke, particulate pollution, chemical vapors, dyes,wood dust, vegetable gums solder and enzymes from foods, molds, fungi,bacteria and their by-products, detergents and pharmaceutical reagents.

In accordance with the present invention, acceptable protocols toadminister IL-12 include the mode of administration and the effectiveamount of IL-12 administered to an animal, including individual dosesize, number of doses and frequency of dose administration.Determination of such protocols can be accomplished by those skilled inthe art. Suitable modes of administration can include, but are notlimited to, oral, nasal, topical, transdermal, rectal, and parenteralroutes. Preferred parenteral routes can include, but are not limited to,subcutaneous, intradermal, intravenous, intramuscular andintraperitoneal routes. Preferred topical routes include inhalation byaerosol (i.e., spraying) or topical surface administration to the skinof an animal.

According to the method of the present invention, an effective amount ofIL-12 to administer to an animal comprises an amount that is capable ofreducing airway hyperresponsiveness (AHR) and/or reducing airflowlimitation and/or symptoms (e.g., shortness of breath, wheezing,dyspnea, exercise limitation or nocturnal awakenings), without beingtoxic to the animal. An amount that is toxic to an animal comprises anyamount that causes damage to the structure or function of an animal(i.e., poisonous). AHR refers to an abnormality of the airways thatallows them to narrow too easily and/or too much in response to astimulus capable of inducing airflow limitation. AHR can be a functionalalteration of the respiratory system caused by inflammation. Airflowlimitation refers to narrowing of airways that can be irreversible orreversible. Airflow limitation can be caused by bronchospasm, airwaysmooth muscle hypertrophy, airway smooth muscle contraction, mucoussecretion, cellular deposits, epithelial destruction, alteration toepithelial permeability, alterations to smooth muscle function orsensitivity, abnormalities of the lung parenchyma and infiltrativediseases in and around the airways.

AHR can be measured by a stress test that comprises measuring ananimal's respiratory system function in response to a provoking agent(i.e., stimulus). AHR can be measured as a change in respiratoryfunction from baseline plotted against the dose of a provoking agent (aprocedure for such measurement is described in detail below in theExamples). Respiratory function can be measured by, for example,spirometry, plethysmographically, peak flows, symptom scores, physicalsigns (i.e., respiratory rate), wheezing, exercise tolerance, use ofrescue medication (i.e., bronchodialators) and blood gases. In humans,spirometry can be used to gauge the change in respiratory function inconjunction with a provoking agent, such as methacholine or histamine.In humans, spirometry is performed by asking a person to take a deepbreath and blow, as long, as hard and as fast as possible into a gaugethat measures airflow and volume. The volume of air expired in the firstsecond is known as forced expiratory volume (FEV₁) and the total amountof air expired is known as the forced vital capacity (FVC). In humans,normal predicted FEV₁ and FVC are available and standardized accordingto weight, height, sex and race. An individual free of disease has anFEV₁ and a FVC of at least about 80% of normal predicted values for aparticular person and a ratio of FEV₁ /FVC of at least about 80%. Valuesare determined before (i.e., representing an animal's resting state) andafter (i.e., representing an animal's higher lung resistance state)inhalation of the provoking agent. The position of the resulting curveindicates the sensitivity of the airways to the provoking agent.

The effect of increasing doses or concentrations of the provoking agenton lung function is determined by measuring the forced expired volume in1 second (FEV₁) and FEV₁ over forced vital capacity (FEV₁ /FVC ratio) ofthe animal challenged with the provoking agent. In humans, the dose orconcentration of a provoking agent (i.e., methacholine or histamine)that causes a 20% fall in FEV₁ (PD₂₀ FEV₁) is indicative of the degreeof AHR. FEV₁ and FVC values can be measured using methods known to thoseof skill in the art.

A variety of provoking agents are useful for measuring AHR values.Suitable provoking agent include direct and indirect stimuli. Preferredprovoking agents include, for example, methacholine (Mch), histamine, anallergen, a leukotriene, saline, hyperventilation, exercise, sulfurdioxide, adenosine, propranolol, cold air, antigen, bradykinin andmixtures thereof. Preferably, Mch is used as a provoking agent.Preferred concentrations of Mch to use in a concentration-response curveare between about 0.001 and about 100 milligram per milliliter (mg/ml).More preferred concentrations of Mch to use in a concentration-responsecurve are between about 0.01 and about 50 mg/ml. Even more preferredconcentrations of Mch to use in a concentration-response curve arebetween about 0.02 and about 25 mg/ml. When Mch is used as a provokingagent, the degree of AHR is defined by the provocative concentration ofMch needed to cause a 20% drop of the FEV₁ of an animal(PC_(20methacholine) FEV₁). For example, in humans and using standardprotocols in the art, a normal person typically has aPC_(20methacholine) FEV₁ >8 mg/ml of Mch. Thus, in humans, AHR isdefined as PC_(20methacholine) FEV₁ <8 mg/ml of Mch.

The effectiveness of a drug to protect an animal from AHR in an animalhaving or susceptible to AHR is measured in doubling amounts. Forexample, the effectiveness an animal to be protected from AHR issignificant if the animal's PC_(2methacholine) FEV₁ is at 1 mg/ml beforeadministration of the drug and is at 2 mg/ml of Mch after administrationof the drug. Similarly, a drug is considered effective if the animal'sPC_(20methacholine) FEV₁ is at 2 mg/ml before administration of the drugand is at 4 mg/ml of Mch after administration of the drug.

In one embodiment of the present invention, an effective amount of IL-12to administer to an animal includes an amount that is capable ofdecreasing methacholine responsiveness without being toxic to theanimal. A preferred effective amount of IL-12 comprises an amount thatis capable of increasing the PC_(20methacholine) FEV₁ of an animaltreated with the interleukin-12 by about one doubling concentrationtowards the PC_(20methacholine) FEV₁ of a normal animal. A normal animalrefers to an animal known not to suffer from or be susceptible toabnormal AHR. A test animal refers to an animal suspected of sufferingfrom or being susceptible to abnormal AHR.

In another embodiment, an effective amount of IL-12 according to themethod of the present invention, comprises an amount that results in animprovement in an animal's PC_(20methacholine) FEV₁ value such that thePC_(20methacholine) FEV₁ value obtained before administration of theIL-12 when the animal is provoked with a first concentration ofmethacholine is the same as the PC_(20methacholine) FEV₁ value obtainedafter administration of the IL-12 when the animal is provoked withdouble the amount of the first concentration of methacholine. Apreferred amount of IL-12 comprises an amount that results in animprovement in an animal's PC_(20methacholine) FEV₁ value such that thePC_(20methacholine) FEV₁ value obtained before administration of theIL-12 is between about 0.01 mg/ml to about 8 mg/ml of methacholine isthe same as the PC_(20methacholine) FEV₁ value obtained afteradministration of the IL-12 is between about 0.02 mg/ml to about 16mg/ml of methacholine.

According to the present invention, respiratory function can beevaluated with a variety of static tests that comprise measuring ananimal's respiratory system function in the absence of a provokingagent. Examples of static tests include, for example, spirometry,plethysmographically, peak flows, symptom scores, physical signs (i.e.,respiratory rate), wheezing, exercise tolerance, use of rescuemedication (i.e., bronchodialators) and blood gases. Evaluatingpulmonary function in static tests can be performed by measuring, forexample, Total Lung Capacity (TLC), Thoracic Gas Volume (TgV),Functional Residual Capacity (FRC), Residual Volume (RV) and SpecificConductance (SGL) for lung volumes, Diffusing Capacity of the Lung forCarbon Monoxide (DLCO), arterial blood gases, including pH, P_(O2) andP_(CO2) for gas exchange. Both FEV₁ and FEV₁ /FVC can be used to measureairflow limitation. If spirometry is used in humans, the FEV₁ of anindividual can be compared to the FEV₁ of predicted values. PredictedFEV₁ values are available for standard normograms based on the animal'sage, sex, weight, height and race. A normal animal typically has an FEV₁at least about 80% of the predicted FEV₁ for the animal. Airflowlimitation results in a FEV₁ less than 80% of predicted values. Analternative method to measure airflow limitation is based on the ratioof FEV₁ and FVC (FEV₁ /FVC). Disease free individuals are defined ashaving a FEV₁ /FVC ratio of at least about 80%. Airflow obstructioncauses the ratio of FEV₁ /FVC to fall to less than 80% of predictedvalues. Thus, an animal having airflow limitation is defined by an FEV₁/FVC less than about 80%.

The effectiveness of a drug to protect an animal having or susceptibleto airflow limitation is determined by measuring the percent improvementin FEV₁ and/or the FEV₁ /FVC ratio before and after administration ofthe drug. In one embodiment, an effective amount of IL-12 comprises anamount that is capable of reducing the airflow limitation of an animalsuch that the FEV₁ /FVC value of the animal is at least about 80%. Inanother embodiment, an effective amount of 12 comprises an amount thatimproves an animal's FEV₁ preferably by between about 6% and about 100%,more preferably by between about 7% and about 100%, and even morepreferably by between about 8% and about 100% of the animal's predictedFEV₁.

It is within the scope of the present invention that a static test canbe performed before or after administration of a provocative agent usedin a stress test.

In another embodiment, an effective amount of IL-12 for use with themethod of the present invention, comprises an amount that is capable ofreducing the airflow limitation of an animal such that the variation ofFEV₁ values of the animal when measured in the evening before bed and inthe morning upon waking is less than about 75%, preferably less thanabout 45%, more preferably less than about 15%, and even more preferablyless than about 8%.

In yet another embodiment, an effective amount of IL-12 for use with themethod of the present invention, comprises an amount that reduces thelevel of IgE in the serum of an animal to between about 0 to about 100international units/ml, preferably between about 10 to about 50international units/ml, more preferably between about 15 to about 25international units/ml, and even more preferably about 20 internationalunits/ml. The concentration of IgE in the serum of an animal can bemeasured using methods known to those of skill in the art. Inparticular, the concentration of IgE in the serum of an animal can bemeasured by, for example, using antibodies that specifically bind to IgEin an enzyme-linked immunoassay or a radioimmunoassay.

In another embodiment, an effective amount of IL-12 for use with themethod of the present invention, comprises an amount that reduceseosinophil blood counts in an animal to preferably between about 0 and470 cells/mm³, more preferably to between about 0 and 300 cells/m³, andeven more preferably to between about 0 and 100 cells/m³. Eosinophilblood counts of an animal can be measured using methods known to thoseof skill in the art. In particular, the eosinophil blood counts of ananimal can be measured by vital stains, such as phloxin B or Diff Quick.

A suitable single dose of IL-12 to administer to an animal is a dosethat is capable of protecting an animal from an inflammatory responsewhen administered one or more times over a suitable time period. Inparticular, a suitable single dose of IL-12 comprises a dose thatimproves AHR by a doubling dose of a provoking agent or improves thestatic respiratory function of an animal. A preferred single dose ofIL-12 comprises between about 1 microgram×kilogram⁻¹ and about 10milligram×kilogram⁻¹ body weight of an animal. A more preferred singledose of IL-12 comprises between about 5 microgram×kilogram⁻¹ and about 7milligram ×kilogram⁻¹ body weight of an animal. An even more preferredsingle dose of IL-12 comprises between about 10 microgram×kilogram⁻¹ andabout 5 milligram×kilogram⁻¹ body weight of an animal. A particularlypreferred single dose of IL-12 comprises between about 0.1milligram×kilogram⁻¹ and about 5 milligram×kilogram⁻¹ body weight of ananimal, if the IL-12 is delivered by aerosol. Another particularlypreferred single dose of IL-12 comprises between about 0.1microgram×kilogram⁻¹ and about 10 microgram ×kilogram⁻¹ body weight ofan animal, if the IL-12 is delivered parenterally.

A preferred IL-12 for use with the method of the present invention,including homologues thereof, is capable of regulating the growth ofnatural killer (NK) and/or T cells. In addition, a preferred IL-12 foruse with the method of the present invention, including homologuesthereof, is capable of increasing antibody derived cytotoxicity orcell-mediated NK cell cytotoxicity. A preferred IL-12 homologue includesat least one epitope capable of regulating the growth of natural killer(NK) and/or T cells substantially similar to that of a natural IL-12counterpart. The ability of an IL-12 homologue to regulate the growth ofNK and/or T cells can be tested using techniques known to those skilledin the art including cellular assays that determine the growth of NKcells or T cells in tissue culture.

In one embodiment, IL-12 suitable for use with the method of the presentinvention comprises IL-12 derived from a substantially similar speciesof animal to which the In-12 is to be administered. For example, ifIL-12 is to be administered to a human patient, then preferably primate,and more preferably human IL-12 can be used to protect the animal from adisease involving inflammation. If IL-12 is to be administered to amouse, then preferably rodent (i.e., hamster, rat, guinea pig), and morepreferably mouse IL-12 can be used. Preferred IL-12 of the presentinvention includes IL-12 derived from primates and rodents. Morepreferred IL-12 includes IL-12 derived from humans and mice, with humanIL-12 being even more preferred.

In another embodiment, IL-12 suitable for use with the method of thepresent invention comprises an isolated protein which has been removedfrom its natural milieu. Isolated IL-12 can, for example, be obtainedfrom its natural source, be produced using recombinant DNA technology,or be synthesized chemically. Preferably, IL-12 of the present inventionincludes recombinantly produced IL-12.

In yet another embodiment, IL-12 suitable for use with the method of thepresent invention comprises IL-12 having sufficient activity tostimulate blast cell proliferation up to half-maximal in a PHAblast cellproliferation assay using RB012892 (J. Exp. Med. 177:1797-1802, 1993).Preferably, IL-12 of the present invention have an activity of fromabout 1×10⁶ to about 6×10⁶ units/mg, more preferably about 2×10⁶ toabout 5×10⁶ units/mg and even more preferably about 4×10⁶ units/mg whenmeasured by the above-referenced blast cell proliferation assay.

In another embodiment, IL-12 of the present invention can beadministered simultaneously or sequentially with an effective amount ofa compound capable of enhancing the ability of the IL-12 to protect ananimal from a disease involving inflammation. The present invention alsoincludes a formulation containing IL-12 and at least one such compoundto protect an animal from a disease involving inflammation. A suitablecompound to be administered simultaneously or sequentially with IL-12includes a compound that is capable of regulating IgE production (i.e.,suppression of interleukin-4 induced IgE synthesis), regulatinginterferon-gamma production, regulating NK cell proliferation andactivation, regulating lymphokine activated killer cells (LAK),regulating T helper cell activity, regulating degranulation of mastcells, protecting sensory nerve endings, regulating eosinophil and/orblast cell activity, preventing or relaxing smooth muscle contraction,reduce microvascular permeability and Th1 and/or Th2 T cell subsetdifferentiation. A preferred compound to be administered simultaneouslyor sequentially with IL-12 includes, but is not limited to, an antigen,an allergen (e.g., plant, such as weed, grass, tree, peanut; animal,such as cat; bacterial; parasitic, such as mite and flea; andmetal-based allergens), a hapten and a drug. A preferred drug to beadministered simultaneously or sequentially with IL-12 includes, but isnot limited to, IL-2, IL-4, anti-IL-4 reagents, glucocorticosteroids,anti-cyclooxygenase agents, anti-cholinergic agents, beta-adrenergicagonists, methyl xanthines (i.e., theophylline), anti-histamines,cromones (i.e., chromoglycate), zyleuton, anti-CD4 reagents, anti-IL-5reagents, surfactants, anti-thromboxane reagents, anti-serotoninreagents, ketotiphen, cytoxin, cylosporin, methotrexate, macrolideantibiotics, troleadomycin, heparin, low molecular weight heparin, andmixtures thereof. The choice of compound to be administered inconjunction with IL-12 can be made by one of skill in the art based onvarious characteristics of the animal. In particular, an animal'sgenetic background, history of occurrence of inflammation, dyspnea,wheezing upon physical exam, symptom scores, physical signs (i.e.,respiratory rate), exercise tolerance, use of rescue medication (i.e.,bronchodialators) and blood gases.

A formulation of the present invention can also include other componentssuch as a pharmaceutically acceptable excipient. For example,formulations of the present invention can be formulated in an excipientthat the animal to be protected can tolerate. Examples of suchexcipients include water, saline, phosphate buffered solutions, Ringer'ssolution, dextrose solution, Hank's solution, polyethyleneglycol-containing physiologically balanced salt solutions, and otheraqueous physiologically balanced salt solutions. Nonaqueous vehicles,such as fixed oils, sesame oil, ethyl oleate, or triglycerides may alsobe used. Other useful formulations include suspensions containingviscosity enhancing agents, such as sodium carboxymethylcellulose,sorbitol, or dextran. Excipients can also contain minor amounts ofadditives, such as substances that enhance isotonicity and chemicalstability or buffers. Examples of buffers include phosphate buffer,bicarbonate buffer and Tris buffer, while examples of preservativesinclude thimerosal, m- or o-cresol, formalin and benzyl alcohol.Standard formulations can either be liquid injectables or solids whichcan be taken up in a suitable liquid as a suspension or solution forinjection. Thus, in a non-liquid formulation, the excipient can comprisedextrose, human serum albumin, preservatives, etc., to which sterilewater or saline can be added prior to administration.

In one embodiment of the present invention, IL-12 or a formulation ofthe present invention can include a controlled release composition thatis capable of slowly releasing the IL-12 or formulation of the presentinvention into an animal. As used herein a controlled releasecomposition comprises IL-12 or a formulation of the present invention ina controlled release vehicle. Suitable controlled release vehiclesinclude, but are not limited to, biocompatible polymers, other polymericmatrices, capsules, microcapsules, microparticles, bolus preparations,osmotic pumps, diffusion devices, liposomes, lipospheres, andtransdermal delivery systems. Other controlled release compositions ofthe present invention include liquids that, upon administration to ananimal, form a solid or a gel in situ. Preferred controlled releasecompositions are biodegradable (i.e., bioerodible).

A preferred controlled release composition of the present invention iscapable of releasing IL-12 or a formulation of the present inventioninto the blood of an animal at a constant rate sufficient to attaintherapeutic dose levels of IL-12 or the formulation to preventinflammation over a period of time ranging from days to months based onIL-12 toxicity parameters. A controlled release formulation of thepresent invention is capable of effecting protection for preferably atleast about 6 hours, more preferably at least about 24 hours, and evenmore preferably for at least about 7 days.

Another embodiment of the present invention comprises a method forprescribing treatment for a respiratory disease involving aninflammatory response, the method comprising: (1) administering to ananimal an effective amount of IL-12; (2) measuring a change in lungfunction in response to a provoking agent in the animal to determine ifthe IL-12 is capable of modulating a disorder selected from the groupconsisting of airway hyperresponsiveness; and (3) prescribing apharmacological therapy effective to reduce inflammation based upon thechanges in lung function. A change in lung function includes measuringstatic respiratory function before and after administration of IL-12. Inaccordance with the present invention, the animal receiving the IL-12 isknown to have a respiratory disease involving inflammation. Measuring achange in lung function in response to a provoking agent can be doneusing a variety of techniques known to those of skill in the art. Inparticular, a change in lung function can be measured by determining theFEV₁, FEV₁ /FVC, PC_(20methacholine) FEV₁ and/or peak flow for therecipient of the provoking agent. Other methods to measure a change inlung function include, for example, lung volumes, peak flows, symptomscores, physical signs (i.e., respiratory rate), wheezing, exercisetolerance, use of rescue medication (i.e., bronchodialators) and bloodgases. A suitable pharmacological therapy effective to reduceinflammation in an animal can be evaluated by determining if and to whatextent the administration of IL-12 has an effect on the lung function ofthe animal. If a change in lung function results from the administrationof IL-12, then that animal can be treated with IL-12. In addition ordepending upon the extent of change in lung function, additionalcompounds can be administered to the animal to enhance the treatment ofthe animal. If no change or a sufficiently small change in lung functionresults from the administration of IL-12, then that animal should betreated with an alternative compound to IL-12. The present method forprescribing treatment for a respiratory disease can also includeevaluating other characteristics of the patient, such as the patient'shistory of respiratory disease, the presence of infectious agents, thepatient's habits (e.g., smoking), the patient's working and livingenvironment, allergies, a history of life threatening respiratoryevents, severity of illness, duration of illness (i.e., acute orchronic), and previous response to other drugs and/or therapy.

Another embodiment of the present invention comprises a method formonitoring the success of a treatment for a respiratory diseaseinvolving an inflammatory response in an animal, the method comprising:(1) administering an effective amount of interleukin-12 to an animalthat has been treated for a respiratory disease involving aninflammatory response; (2) measuring a change in the lung function ofthe animal in response to a provoking agent of the present invention;and (3) monitoring the success of the treatment by comparing the changein lung function with previous measurements of lung function in theanimal. If the treatment does not result in the improvement of lungfunction, then the administration of IL-12 should be able to alter lungfunction Conversely, if the treatment does result in lung functionimprovement, then the administration of IL-12 should not alter lungfunction because the lung function will have been improved by theoriginal treatment. The monitoring of success can also include comparingthe change in lung function before and after administration of IL-12 toan animal with other aforementioned characteristics of the animal.

Another embodiment of the present invention includes a method forlong-term care of a patient having a disease involving inflammation, themethod comprising: (1) assessing the status of the disease of a patient;(2) administering to the patient an effective amount of IL-12; and (3)providing long-term care of the patient by preventing significantexposure of the patient to the cause of the disease. Preferably, thestatus of the disease is assessed by determining a characteristic of thedisease, such as determining the form, severity and complications of thedisease. In addition, the status of the disease is assessed bydetermining, for example, a causative agent and/or a provoking agent ofthe disease. From the assessment of the causative and/or provoking agentof the disease, long-term care can be provided by minimizing theexposure of the patient to the causative and/or provoking agent of thedisease.

The following examples are provided for the purposes of illustration andare not intended to limit the scope of the present invention.

EXAMPLES Example 1

This example describes the effect of administration IL-12 to BALB/c miceimmunized with ovalbumin using alum as an adjuvant.

A. Experiment 1

Airway hyperresponsiveness was assessed in mice (i.e., stress tests) inthe following manner. Six to 8 week old BALB/c female mice, weighingabout 18-25 grams, were obtained from Jackson Laboratories (Bar Harbor,Minn.). The mice were quarantined for 2-3 weeks. The mice were then bledand the blood was assayed for the presence of antibody to common murineviruses (e.g., Minute virus, hepatitis virus, mycoplasma pulmonis,pneumonia virus and sendai virus) using methods standard in the art.After the mice were established to be virus free, the mice were housedon cedar chips (J. P. Murphy Forest Products, Montville, N.J.), in microisolators (Allentown Caging Equipment, Co., Allentown, N.J.) placed inHEPA flow hoods (NuAire Animal Isolators, Plymouth, Minn.) usingstandard barrier techniques. The mice were allowed free access to waterand fed Purina's 5L31 Pico Rodent Chow (Ralston Purina, St. Louis, Mo.)ab Yibra.

Eleven mice (IP/OVA) were injected intraperitoneally with about 10 μgovalbumin (OVA) (Sigma, St. Louis, Mo.) in 100 μg aluminum hydroxide(Alum) (T. J. Baker Chemical Co., Phillipsburg, N.J.) dissolved inphosphate buffered saline (PBS). Five mice that were housed underidentical conditions as described above did not receive any treatmentsand are referred to herein as non-immune (NIM) mice.

Two weeks after priming, the 11 OVA injected mice were exposed to a 1%aerosolized solution of OVA in PBS for 30 minutes daily for 8 days(i.e., 8 exposures in 8 days) and are referred to herein as IP/OVA+AERO.Six of the 11 mice were treated with 1 μg of murine recombinant IL-12intraperitoneally (IP) one hour before each of the 8 daily aerosolizedOVA treatments.

Twenty four hours following the last aerosol exposure, the mice wereattached to the following equipment to measure the respiratory functionof each mouse when challenged with a provoking agent. The mice wereanesthetized with 70 mg/kg of intraperitoneal pentobarbital sodium(Abbott Laboratories, Chicago, Ill.) and the trachea and right internaljugular vein were exposed. A metal 19 gauge endotracheal catheter wasinserted and sutured into the trachea, and a 0.0048 cm internal diameter×5 cm Silastic catheter (Dow Corning Corp., Midland, Mich.) was insertedand sutured into the right internal vein. Following surgery, the micewere in a plethysmographic chamber and the tracheostomy tube wasattached to a 4-way connector (Small Parts, Inc., Miami Lakes, Fla.),with one port connected to a catheter measuring airway opening pressure(P_(AO)) and two ports connected to the inspiratory and expiratory portsof a volume cycled ventilator (Harvard Apparatus Rodent Ventilator,Model 680, South Natwick, Mass.). The mice were ventilated at 200breaths per minute, tidal volume of 5-6 ml/kg, and with 2 cm H₂ Opositive end-expiratory pressure. Adequacy of alveolar ventilation wasconfirmed by the lack of spontaneous respiration (i.e., over-breathing),and transcutaneous CO_(s) measurements. Transpulmonary pressure wasestimated as the P_(AO), referenced to pressure within theplethysmographic using a differential pressure transducer (ValidyneModel MP-45-1-871, Validyne Engineering Corp., Northridge, Calif.).Changes in volume were determined by pressure changes in theplethysmographic chamber referenced to pressure in a reference box usinga second differential pressure transducer. The two transducers andamplifiers were electronically phased to less than 5 degrees from 1 to30 Hz and then converted from an analog to digital signal using a 16 bitanalog to digital board Model NB-MIO-16X-18 (National Instruments Corp.,Austin, Tex.) at 600 bits per second per channel. The digitized signalswere fed into a Macintosh Quadra 800 computer (Model M1206, AppleComputer, Inc., Cupertino, Calif.) and analyzed using the real timecomputer program LabVIEW (National Instruments Corp., Austin, Tex.).Flow was determined by differentiation of the volume signal andcompliance was calculated as the change in volume divided by the changein pressure at zero flow points for the inspiratory phase and expiratoryphase. Average compliance was calculated as the arithmetic mean ofinspiratory and expiratory compliance for each breath. The LabVIEWcomputer program used pressure, flow, volume and average compliance tocalculate pulmonary resistance (R1) and compliance according to themethod of Amdur et al. (pp. 364-368, 1958, Am. J. Physiol., vol. 192).The breath by breath results for Rl, compliance, conductance andspecific compliance were tabulated and the reported values are theaverage of at least 10-20 breaths at the peak of response for each dose.

Following placement in a plethysmographic chamber, each mouse waschallenged with methacholine to assess airway hyperresponsive pulmonaryfunction. In vivo airway hyperresponsiveness (AHR) was assessed as thechange in respiratory system function after noncumulative, intravenousmethacholine (i.e., Acetyl-β-methylcholine) challenge (McDRC).Acetyl-β-methylcholine (Aldrich Chemical, Milwaukee, Wis.) was dissolvedin normal saline and administered into the internal jugular veincatheter with a micro syringe (Hamilton, Co., Reno, Nev.). AHR wasassessed as the resistance (Rl) in cmH₂ O/ml/sec followingadministration of 6 tripling doses of about 5 μg/mg to about 1233 μg/mgof intravenous methacholine.

The means and standard errors of the log 10 of resistance (Rl) by doseof methacholine and by group obtained from the stress test areillustrated in FIG. 1 (n=the number of mice in each treatment group).The results indicate that administration of IL-12 to the OVA sensitizedmice significantly blocks airway hyperresponsiveness to methacholinechallenge. It should be noted that measuring the Rl value in a mouse,can be used to diagnose airflow obstruction similar to measuring theFEV_(l) and/or FEV₁ /FVC ratio in a human.

B. Experiment 2

A separate set of mice were prepared as described above in Section A buton a different day. Eight mice were injected IP with OVA and treatedwith 1% aerosolized solution of OVA in PBS as described in section A.Five non-immune mice were prepared as described in section A. The 8OVA-treated mice were challenged with methacholine using the methoddescribed above in Section A. The results of this second experiment areillustrated in FIG. 2 and are statistically identical to the results ofExperiment 1 (Section A), thereby confirming the results of Experiment1.

C. Combined Analysis

A statistical analysis of the data was performed on the log 10 ofresistance using Akaike Information Criteria repeated analysis variance.The repeated measures analysis of variance model with the unstructuredvariance matrix and fixed effects for group, dose and interaction ofgroup by dose resulted in a significant interaction of p=0.0001. Foreach dose of methacholine, analysis of variance (ANOVA) was performedallowing for unequal variances. Pairwise comparisons were performed onlyif the ANOVA was significant for the given dose. The procedure used isan extension of Fisher's protected least significant difference multiplecomparison procedure. Rl value for baseline, saline 1, saline 2 and eachdose of methacholine ±the standard error of the mean (SEM) are shownbelow in Table 1. The statistical values summarized in Table 1 arerepresented in FIG. 3.

                                      TABLE I                                     __________________________________________________________________________    MEANS AND STANDARD ERROR OF THE LOG MEAN RESISTANCE (CM.sub.H2O/ML/SEC)       BY IV METHACHOLINE (μg/gram) Dose of IV METHACHOLINE (μg/mg)                                 Baseline                                                                           5.08                                                                              15.23                                                                             45.68                                                                             137.04                                                                            411.07                                                                            1233.21                         __________________________________________________________________________    1   NIM       MEAN   -.013                                                                              -0.14                                                                             -0.11                                                                             0.01                                                                              0.40                                                                              1.26                                                                              1.79                                          STDERR 0.05 0.05                                                                              0.05                                                                              0.05                                                                              0.07                                                                              0.09                                                                              0.12                                          N      10   10  10  10  10  10  10                              2   IP/OVA + AERO                                                                           MEAN   -0.09                                                                              -0.09                                                                             -0.04                                                                             0.18                                                                              1.14                                                                              2.48                                                                              3.10                                          STDERR 0.04 0.04                                                                              0.04                                                                              0.05                                                                              0.19                                                                              0.20                                                                              0.12                                          N      10   10  10  10  10  10  10                              3   IP/OVA + AERO                                                                           MEAN   -0.11                                                                              -0.11                                                                             -0.08                                                                             0.05                                                                              0.62                                                                              1.51                                                                              2.20                                +IL-12    STDERR 0.03 0.03                                                                              0.02                                                                              0.04                                                                              0.11                                                                              0.15                                                                              0.13                                          N      9    9   9   9   9   9   9                               SIGNIFICANT CONTRASTS BY REPEATED                                                                  NONE NONE                                                                              NONE                                                                              2 vs 1                                                                            2 vs 1                                                                            2 vs 1                                                                            2 vs 1                          ANALYSIS OF VARIANCE USING AKAIKE     2 vs 3                                                                            2 vs 3                                                                            2 vs 3                          INFORMATION CRITERIA                          1 vs 3                          __________________________________________________________________________     N = number of mice/group                                                      Saline 1 and Saline 2 not included                                       

The results indicate that the pairwise comparison listed in the AkaikeInformation Criteria analysis were found to be significant, p<0.05. Theoverall p value using repeated analysis of variance is highlysignificant at p=0.0001. The results also indicate that over a 5-foldincrease in AHR to methacholine following OVA immunization in BALB/cmice occurred. Administration of IL-12 blocks about 80% of the increasein 0VA-induced AHR. Thus, IL-12 is a potent inhibitor of antigen inducedAHR.

While various embodiments of the present invention have been describedin detail, it is apparent that modifications and adaptations of thoseembodiments will occur to those skilled in the art. It is to beexpressly understood, however, that such modifications and adaptationsare within the scope of the present invention, as set forth in thefollowing claims.

What is claimed:
 1. A method to protect a mammal from a respiratorydisease involving an inflammatory response, wherein said airwayhyperresponsiveness and/or airflow limitation associated with arespiratory disease involving an inflammatory response, said methodcomprising administering to said mammal an effective amount ofinterleukin-12.
 2. The method of claim 1, wherein said disease comprisesa chronic obstructive pulmonary disease of the airways.
 3. The method ofclaim 1, wherein said disease is selected from the group consisting ofasthma, allergic bronchopulmonary aspergillosis, hypersensitivitypneumonia, eosinophilic pneumonia, emphysema, bronchitis, allergicbronchitis bronchiectasis, cystic fibrosis, hypersensitivitypneumotitis, occupational asthma, sarcoid, reactive airway diseasesyndrome, interstitial lung disease, hyper-eosinophilic syndrome andparasitic lung disease.
 4. The method of claim 1, wherein said diseaseis selected from the group consisting of asthma, occupational asthma andreactive airway disease syndrome.
 5. The method of claim 1, wherein saidinterleukin-12 is administered by at least one route selected from thegroup consisting of oral, nasal, topical, inhaled, transdermal, rectaland parenteral routes.
 6. The method of claim 1, wherein saidinterleukin-12 is administered by at least one route selected from thegroup consisting of inhalation, intramuscular injection and subcutaneousinjection.
 7. The method of claim 1, wherein said interleukin-12 isadministered by inhalation.
 8. The method of claim 1, wherein aneffective amount of said interleukin-12 comprises an amount that iscapable of reducing airway hyperresponsiveness without being toxic tosaid mammal.
 9. The method of claim 1, wherein an effective amount ofsaid interleukin-12 comprises an amount that is capable of decreasingmethacholine responsiveness without being toxic to said mammal.
 10. Themethod of claim 1, wherein an effective amount of said interleukin-12,comprises an amount that results in an improvement in a mammal'sPC_(20methacholine) FEV₁ value such that the PC_(20methacholine) FEV₁value obtained before administration of the IL-12 when the mammal isprovoked with a first concentration of methacholine is the same as thePC_(20methacholine) FEV₁ value obtained after administration of theIL-12 when the mammal is provoked with double the amount of the firstconcentration of methacholine.
 11. The method of claim 12, wherein saidfirst concentration of methacholine is between about 0.01 mg/ml andabout 8 mg/ml.
 12. The method of claim 1, wherein an effective amount ofsaid interleukin-12 comprises an amount that improves an animal's FEV₁by between about 6% and about 100% of said animal's predicted FEV₁. 13.The method of claim 1, of said interleukin-12 comprises an amount thatimproves a mammal's Fev₁ by between about 7% and about 100% of saidanimal's predicted FEV₁.
 14. The method of claim 1, wherein an effectiveamount of said interleukin-12 comprises an amount that improves amammal's FEV₁ by between about 8% and about 100% of said mammal'spredicted FEV₁.
 15. The method of claim 1, wherein an effective amountof said interleukin-12 comprises an amount that is capable of reducingthe airflow limitation of a mammal's such that the FEV₁ /FVC value ofsaid mammal's is at least about 80%.
 16. The method of claim 1, whereinan effective amount of said interleukin-12 comprises an amount thatreduces the level of IgE in the serum of a mammal's to between about 0to about 100 international units/ml.
 17. The method of claim 1, whereinan effective amount of said interleukin-12 comprises an amount thatreduces the level of IgE in a mammal to between about 10 to about 50international units/ml.
 18. The method of claim 1, wherein an effectiveamount of said interleukin-12 comprises an amount that reduces the levelof IgE in the serum of a mammal to between about 15 to about 25international units/ml.
 19. The method of claim 1, wherein an effectiveamount of said interleukin-12 comprises an amount that reduces the levelof IgE in the serum of a mammal about 20 international units/ml.
 20. Themethod of claim 1, wherein an effective amount of said interleukin-12comprises an amount that reduces eosinophil blood counts to in a mammalto between about 0 and 470 cells/mm³.
 21. The method of claim. 1,wherein an effective amount of said interleukin-12 comprises an amountthat reduces eosinophil blood counts in a mammal to between about 0 and300 cells/mm³.
 22. The method of claim 1, wherein an effective amount ofsaid interleukin-12 comprises an amount that reduces eosinophil bloodcounts in a mammal to between about 0 and 100 cells/mm³.
 23. The methodof claim 1, wherein an effective amount of said interleukin-12 comprisesbetween about 1 microgram×kilogram⁻¹ and about 10 milligram ×kilogram⁻¹body weight of a mammal.
 24. The method of claim 1, wherein an effectiveamount of said interleukin-12 comprises between about 5 microgram×kilogram⁻¹ and about 7 milligram×kilogram⁻¹ body weight of a mammal.25. The method of claim 1, wherein an effective amount of saidinterleukin-12 comprises between about 10 microgram ×kilogram⁻¹ andabout 5 milligram ×kilogram⁻¹ body weight of a mammal.
 26. The method ofclaim 1, wherein an effective amount of said interleukin-12 comprisesbetween about 0.1 milligram ×kilogram⁻¹ and about 5 milligram×kilogram⁻¹body weight of a mammal, if said interleukin-12 is delivered by aerosol.27. The method of claim 1, wherein an effective amount of saidinterleukin-12 comprises between about 0.1 microgram ×kilogram⁻¹ andabout 10 microgram ×kilogram⁻¹ body weight of an mammal, if saidinterleukin-12 is delivered parenterally.
 28. The method of claim 1,wherein said interleukin-12 has an activity of from about 1×10⁶ to about6×10⁶ units/milligram.
 29. The method of claim 1, wherein saidinterleukin-12 has an activity of from about 2×10⁶ to about 5×10⁶units/milligram.
 30. The method of claim 1, wherein said interleukin-12has an activity of about 4×10⁶ units/milligram.
 31. The method of claim1, wherein said interleukin-12 is produced recombinantly.
 32. The methodof claim 1, wherein said interleukin-12 is selected from a groupconsisting of murine and human interleukin-12.
 33. The method of claim1, wherein said interleukin-12 is human interleukin-12.
 34. The methodof claim 1, wherein said mammal is selected from the group consisting ofhumans, cats, rodents and horses.
 35. The method of claim 1, furthercomprising administering simultaneously or sequentially with said IL-12,an effective amount of an antigen.
 36. The method of claim 1, whereinsaid interleukin-12 is administered in a pharmaceutically acceptableexcipient.
 37. The method of claim 36, wherein said pharmaceuticallyacceptable excipient comprises a slow release vehicle.
 38. A method forprescribing treatment for airway hyperresponsiveness and/or airflowlimitation associated with a respiratory disease involving aninflammatory response, comprising:a. administering to a mammal aneffective amount of interleukin-12; b. measuring a change in lungfunction in response to a provoking agent in said mammal to determine ifsaid interleukin-12 is capable of modulating airway hyperresponsiveness;and c. prescribing a pharmacological therapy comprising administrationof IL-12 to said mammal effective to reduce inflammation based upon saidchanges in lung function.
 39. The method of claim 38, wherein saidprovoking agent is selected from the group consisting of a direct and anindirect stimuli.
 40. The method of claim 38, wherein said provokingagent is selected from the group consisting of an allergen,methacholine, histamine, a leukotriene, saline, hyperventilation,exercise, sulfur dioxide, adenosine, propranolol, cold air, antigen,bradykinin and mixtures thereof.
 41. The method of claim 38, whereinsaid step of measuring comprises measuring a value selected from thegroup. Consisting of FEV₁, FEV₁ /FVC, PC_(20methacholine) FEV₁ and peakflow.
 42. A method for monitoring the success of a treatment for in amammal, airway hyperresponsiveness and/or airflow limitation associatedwith a respiratory disease involving a inflammatory response, saidmethod comprising:a. administering an effective amount of interleukin-12to a mammal that has been treated for a respiratory disease involving aninflammatory response; b. measuring a change in lung function in saidmammal in response to a provoking agent; and c. monitoring the successof said treatment by comparing said change in lung function withprevious measurements of lung function in said mammal.
 43. The method ofclaim 42, wherein said treatment comprises administering a therapeuticreagent selected from the group consisting of IL-2, IL-4,glucocorticosteroids, anti-cyclooxygenase agents, anti-cholinergicagents, beta-adrenergic agonists, methyl xanthines, anti-histamines,cromones, zyleuton, ketotiphen, cytoxin, cylosporin, methotrexate,macrolide antibiotics, troleadomycin, heparin, low molecular weightheparin, and mixtures thereof.